QuantiTect Primer Assays are genomewide, bioinformatically validated primer sets for use in SYBR Green-based real-time RT-PCR on any cycler. Assays are available for all genes from human, rat, mouse, and many other species. Each assay for a specific gene is supplied as a lyophilized mix of forward and reverse primers that can be easily reconstituted to obtain a 10x assay solution (reaction components for real-time RT-PCR need to be ordered separately). When used in combination with QuantiFast, QuantiTect, Rotor-Gene, or FastLane Kits for SYBR Green detection, QuantiTect Primer Assays guarantee highly specific and sensitive results in real-time RT-PCR that are comparable to probe-based detection.

Search in GeneGlobe

You can search for the following terms:

Entrez Gene IDs (e.g., 835)

RefSeq IDs (e.g., NM_032983, NP_116765)

Gene symbols (e.g., CASP2)

Cat. no. (e.g., SI00299551, QT01342509)

Sanger ID or Accession (e.g., hsa-let-7b, MI0000063)

CpG loci identification numbers (CG#) (e.g., CG17753661)

Do not enter species information in the Search box. Use the drop-down list to select your species of interest.
When searching for miRNAs, do not omit the hyphens. Use hyphens or spaces (e.g., search for hsa-let-7b or hsa let 7b. Do not search for hsalet7b).

QuantiTect Primer Assays are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.

Superior sensitivity in real-time RT-PCR.

MAPK1 (a protein kinase) and IL6 (a cytokine) in human leukocyte cDNA (1 ng) were quantified in duplicate using [A] QuantiTect Primer Assays and the QuantiTect SYBR® Green PCR Kit, or [B] probe-based assays and a real-time PCR kit from Supplier AII. QuantiTect Primer Assays provided greater sensitivity than the probe-based assays, as demonstrated by the lower CT values, as well as high specificity, as indicated by the single peaks in melting curve analysis.

Reproducible real-time RT-PCR.

[A] Different amounts of leukocyte cDNA were analyzed in duplicate using the QuantiTect Primer Assay for IL8 (a chemokine) and QuantiFast SYBR® Green PCR Kit. No template control (NTC) reactions were also performed (shown by the flat plots). [B] Melting curve analysis, demonstrating high PCR specificity.

QuantiTect Primer Assays enable reproducible quantification of RNA transcripts over a range of template amounts (see part [A] of figure "Reproducible real-time RT-PCR"). Accurate quantification is guaranteed when the assays are run using QuantiFast SYBR Green Kits or other SYBR Green kits from QIAGEN. The kits prevent the formation of PCR artifacts (see part [B] of figure "Reproducible real-time RT-PCR").

When used with QuantiFast, QuantiTect, Rotor-Gene, or FastLane SYBR Green Kits (master mixes for two-step or one-step RT-PCR), the assays may even give lower CT values compared with probe-based assays (see figure "Superior sensitivity in real-time RT-PCR"). This is due to the high PCR specificity of the kits and multiple binding of SYBR Green dye to each PCR product. Highly sensitive quantification is possible even with low-abundance transcripts.

Principle

QuantiTect Primer Assays eliminate the need to develop and validate real-time RT-PCR assays (see figure "Time savings in development of real-time RT-PCR assays"). They can be used in both one-step and two-step qRT-PCR. Simply visit QIAGEN's GeneGlobe Web portal, and select and order an assay for any gene from human, rat, mouse, Drosophila, Arabidopsis, and other species. QuantiTect Primer Assays provide guaranteed results when used with QuantiFast, QuantiTect, Rotor-Gene, or FastLane SYBR Green Kits (master mixes for two-step or one-step RT-PCR).

Procedure

Each QuantiTect Primer Assay is supplied as a lyophilized mix of forward and reverse primers for a specific gene. The primers are reconstituted in TE, pH 8.0 to give a 10x primer solution, which is then added to the master mix supplied with a QuantiFast, QuantiTect, Rotor-Gene, or FastLane Kit for SYBR Green detection. The master mix with primers is aliquoted into PCR tubes or wells, and the individual template samples are then added.

QuantiTect Primer Assays provide guaranteed results in gene expression analysis when used in combination with one of the following kits shown in the table.

Two-step and one-step qRT-PCR kits for use with QuantiTect Primer Assays

Two-step qRT-PCR

One-step qRT-PCR

QuantiFast SYBR Green PCR Kit

QuantiFast SYBR Green RT-PCR Kit

Rotor-Gene SYBR Green PCR Kit

Rotor-Gene SYBR Green RT-PCR Kit

QuantiTect SYBR Green PCR Kit

QuantiTect SYBR Green RT-PCR Kit

QuantiTect Primer Assays are designed to detect RNA only where possible. For assays where detection of genomic DNA cannot be avoided, cDNA can be prepared using the QuantiTect Reverse Transcription Kit or the FastLane Cell cDNA Kit. Both kits provide cDNA synthesis with integrated removal of genomic DNA contamination.

Naming

Each QuantiTect Primer Assay available for a particular gene is given a number that appears in the product name after the gene symbol. Read more in the GeneGlobe FAQs.

Applications

QuantiTect Primer Assays are for gene expression analysis using one-step and two-step qRT-PCR with SYBR Green detection. They are highly suited for applications such as: